The following includes information that may be useful in understanding the present inventions. It is not an admission that any of the information provided herein is prior art, or relevant, to the presently described or claimed inventions, or that any publication or document that is specifically or implicitly referenced is prior art.
In humans and other mammals wound injury triggers an organized complex cascade of cellular and biochemical events that will in most cases result in a healed wound. An ideally healed wound is one that restores normal anatomical structure, function, and appearance on cellular, tissue, organ, and organism levels. Wound healing, whether initiated by trauma, microbes or foreign materials, proceeds via a complex process encompassing a number of overlapping phases, including inflammation, epithelialization, angiogenesis and matrix deposition. Normally, these processes lead to a mature wound and a certain degree of scar formation. Although inflammation and repair mostly occur along a prescribed course, the sensitivity of the process is dependent on the balance of a variety of wound healing molecules, including for example, a network of regulatory cytokines and growth factors.
Wounds that do not heal at normal or expected rates, including chronic wounds, such as diabetic foot ulcers, pressure ulcers, and venous leg ulcers (VLU), are an increasing worldwide problem. It is estimated, for example, that 1-2% of the population in Western countries will develop a chronic wound over the course of their lifetimes. Chronic wounds represent a major economic burden on healthcare services, with an estimated annual expenditure in the United States alone of up to $25 billion. Kuehn B M, Chronic wound care guidelines issued. JAMA 297: 938-939, 2007. An estimated 1.3 million to 3 million US individuals are believed to have pressure ulcers; and as many as 10-15% of the 20 million individuals with diabetes are at risk of developing chronic ulcers. Many more have venous ulcers or wounds that result from arterial disease. With growing numbers of elderly and diabetics in the population, this expenditure figure is expected to rise in coming years. Unfortunately, there are few effective therapeutic options for these debilitating wounds, and there remains a significant need for effective new treatments. While, over the years, basic and clinical research has revealed much about the individual molecular and cellular processes involved in wound healing, attempts to accelerate and/or improve wound healing by enhancing, inhibiting, or modifying isolated aspects of the wound healing process have met with only limited success.
Scars are the result of wounds that have healed, lesions due to diseases, or surgical operations. Hypertrophic and keloid scars occur when the tissue response is out of proportion to the amount of scar tissue required for normal repair and healing. Certain regions of the body, including back, shoulders, sternum and earlobe, are especially prone to develop abnormal scars known as hypertrophic scars or keloids. These scars are bulky lesions representing an increased deposition of collagen fibers. They have the same clinical appearance: they are red, raised, and firm and posses a smooth, shiny surface. Whereas hypertrophic scars can flatten spontaneously in the course of one to several years, keloids persist and extend beyond the site of the original injury. As thickened red scars that exceed the boundary of an injury and may grow for a prolonged period of time, keloids are hyperplastic connective tissue masses that occur in the dermis and adjacent subcutaneous tissue, most commonly following trauma, in certain susceptible individuals. Keloid lesions are formed when local skin fibroblasts undergo vigorous hyperplasia and proliferation in response to local stimuli. The increase in scar size is due to deposition of increased amounts of collagen into the tissue. African-Americans are genetically prone to developing keloids. Keloid development has been associated with different types of skin injury including surgery, ear piercing, laceration, burns, vaccination or inflammatory process. Hypertrophic scars are masses which can result from burns or other injuries to the skin. Such scars are usually permanent and resistant to known methods of therapy. Patients suffering from hypertrophic scars or keloids complain about local pain, itchiness and local sensitivity, all of which compromise their quality of life as well as affect the individual body image.
Various therapies for keloids have had only limited success. Existing efforts to manage hypertrophic scars and keloids include surgery, mechanical pressure, steroids, x-ray irradiation and cryotherapy. Disadvantages have been reported to be associated with each of these methods. For example, surgical removal of the scar tissue may be often incomplete and can result in the development of hypertrophic scars and keloids at the incision and suture points, i.e., scarring frequently recurs after a keloid is surgically removed, and steroid treatments may be unpredictable and often result in depigmentation of the skin. Simple surgical excision of keloid scars has a 50%-80% risk of recurrence. A combination of surgery with either intralesional corticosteroid injection or radiotherapy has been a typical treatment. However, intralesional corticosteroid injection is prone to complications (fat atrophy, dermal thinning, and pigment changes).
Atrophic or depressed scars resulting from an inflammatory episode are characterized by contractions of the skin, and leave a cosmetically displeasing and permanent scar. The most common example is scarring which occurs following inflammatory acne or chickenpox. The depression occurs as a normal consequence of wound healing, and the scar tissue causing the depression is predominantly comprised of collagen resulting from fibroblast proliferation and metabolism. Some acne patients are successfully treated using steroids injected intralesionally, topical liquid nitrogen applications, or dermabrasion. In many cases, however, there is either no improvement or the treatment results in other complications.
Scars that cross joints or skin creases at right angles are prone to develop shortening or contracture. Scar contractures occur when the scar is not fully matured, often tend to be hypertrophic, and are typically disabling and dysfunctional. They are common after burn injury across joints or skin concavities. For scar contractures, surgical release with splinting, acrylic casting, and compression therapy may be required. Full thickness and split or partial thickness skin grafts and, perhaps more effectively, local and free flaps are used for reconstruction of difficult and extensive scars and contractures.
Adhesion formation is a process in which bodily tissues that are normally separate become connected by scar tissue. Adhesions most commonly result from surgical incision, abrasion, or trauma. Adhesions can form following most any type of surgery, but develop with the highest frequency following general abdominal, gynecologic, orthopedic, and cardiac surgeries. It has been reported that following abdominal surgery the incidence of peritoneal adhesion formation may be as high as 90%. See U.S. Pat. No. 6,613,325. The incidence of adhesion formation is also thought to be as high as 90% in patients that have undergone multiple surgeries. Post operative intraperitoneal and pelvic adhesions represent a major problem in patients recovering from surgery in the abdominal cavity, where there is a tendency for adhesions to form between the affected tissues. See U.S. Pat. No. 5,002,551. The pervasiveness of this problem also has severe economic consequences.
Although adhesions occur most commonly following surgery, adhesions may also occur from tissue damage other than surgery, including traumatic injury, inflammatory disease, intraperitoneal chemotherapy and radiation therapy. Amongst other complications, the presence of surgical adhesions may be associated with pain, discomfort, and female infertility resulting from gynecological surgery. Intestinal obstructions, for example, are a complication that results from surgical adhesions. Adhesions are also reported to be a leading cause of bowel obstruction and infertility, and related complications include chronic pelvic pain, urethral obstruction and voiding dysfunction. See U.S. Pat. No. 6,689,803. Adhesion formation may result from injury to the peritoneum, which in turn may cause the site of injury or trauma to become inflamed. Although inflammation is a part of the healing process, it can contribute to adhesion formation by contributing to the development of fibrous bands of scar tissue. Through a process called fibrinolysis, the fibrin bands eventually dissolve. However, where fibrin bands do not dissolve, they can develop into proliferating adhesions that connect and bind to organs and tissues that are normally separate. It has been reported that excess production and deposition of the extracellular matrix may be a key factor in producing tissue fibrosis throughout the body including the development of peritoneal adhesions (see U.S. Pat. No. 6,841,153).
Various approaches for the prevention of adhesion formation have been reported. See Dizerega, G. S. & Rodgers, K. E., “Prevention of Postoperative Adhesions,” in “The Peritoneum,” Dizerega, G. S. & Rodgers, K. E., eds., Springer-Verlang, New York, pp. 307-369 (1992). General categories of treatment for adhesions that have been reported, include: 1) prevention of fibrin deposition in the peritoneal exudate, 2) reduction of local tissue inflammation; and 3) removal of fibrin deposits. Id. However, despite years of research it has been reported that very few products for the prevention of post-operative adhesions have resulted. Johns, A., Human Reproductive Update, 7 (6):577-579 (2001). Meanwhile, the medical problems associated with surgical adhesions are becoming more serious because there is a general rise in repeat surgical procedures for a number of disorders. Thus, there is a vital need for the development of compounds and methods for preventing surgical adhesions and mitigating the complications they cause.
Fibroproliferative diseases, including pulmonary fibrosis, systemic sclerosis, liver cirrhosis, cardiovascular disease, progressive kidney disease, and macular degeneration, are a leading cause of morbidity and mortality and can affect all tissues and organ systems. Fibrotic tissue remodeling can also influence cancer metastasis and accelerate chronic graft rejection in transplant recipients. Nevertheless, despite its enormous impact on human health, there are currently no approved treatments that directly target the mechanism(s) of fibrosis.
Fibrosis is the abnormal accumulation of fibrous tissue that can occur as a part of the wound-healing process in damaged tissue. Examples of fibrosis include liver fibrosis, lung fibrosis (e.g., silicosis, asbestosis, idiopathic pulmonary fibrosis), oral fibrosis, endomyocardial fibrosis, retroperitoneal fibrosis, deltoid fibrosis, kidney fibrosis (including diabetic nephropathy), and glomerulosclerosis. Liver fibrosis, for example, occurs as a part of the wound-healing response to chronic liver injury. Fibrosis can occur as a complication of haemochromatosis, Wilson's disease, alcoholism, schistosomiasis, viral hepatitis, bile duct obstruction, exposure to toxins, and metabolic disorders. This formation of fibrotic tissue is believed to represent an attempt by the body to encapsulate injured tissue. Liver fibrosis is characterized by the accumulation of extracellular matrix that can be distinguished qualitatively from that in normal liver. Left unchecked, hepatic fibrosis progresses to cirrhosis (defined by the presence of encapsulated nodules), liver failure, and death. Endomyocardial fibrosis is an idiopathic disorder that is characterized by the development of restrictive cardiomyopathy. In endomyocardial fibrosis, the underlying process produces patchy fibrosis of the endocardial surface of the heart, leading to reduced compliance and, ultimately, restrictive physiology as the endomyocardial surface becomes more generally involved. Endocardial fibrosis principally involves the inflow tracts of the right and left ventricles and may affect the atrioventricular valves, leading to tricuspid and mitral regurgitation. Oral submucous fibrosis is a chronic, debilitating disease of the oral cavity characterized by inflammation and progressive fibrosis of the submucosal tissues (lamina propria and deeper connective tissues). It results in marked rigidity and an eventual inability to open the mouth. The buccal mucosa is the most commonly involved site, but any part of the oral cavity can be involved, even the pharynx. Retroperitoneal fibrosis is characterized by the development of extensive fibrosis throughout the retroperitoneum, typically centered over the anterior surface of the fourth and fifth lumbar vertebrae. This fibrosis leads to entrapment and obstruction of retroperitoneal structures, notably the ureters. In most cases, the etiology is unknown. However, its occasional association with autoimmune diseases and its response to corticosteroids and immunosuppressive therapy suggest it may be immunologically mediated. Deltoid fibrosis is a muscle disorder marked by intramuscular fibrous bands within the substance of the deltoid muscle. These bands lead to secondary contractures that affect the function of the shoulder joint. Scapular winging and secondary scoliosis also may be related to this condition. Deltoid fibrosis has been associated with fibrous contractures of the gluteal and quadriceps muscles and is likely a similar process
Understanding of the cellular and biochemical mechanisms underlying liver fibrosis has advanced in recent years (reviewed by Li and Friedman, J. Gastroenterol. Hepatol. 14:618-633, 1999). Stellate cells are believed to be a major source of extracellular matrix in the liver. Stellate cells respond to a variety of cytokines present in the liver, some of which they also produce (Friedman, Seminars in Liver Disease 19:129-140, 1999). As summarized by Li and Friedman, actual and proposed therapeutic strategies for liver fibrosis include removal of the underlying cause (e.g., toxin or infectious agent), suppression of inflammation (using, e.g., corticosteroids, IL-1 receptor antagonists, or other agents that may suppress inflammation), down-regulation of stellate cell activation (using, e.g., gamma interferon or antioxidants), promotion of matrix degradation, or promotion of stellate cell apoptosis. Despite recent progress, many of these strategies are still in the experimental stage, and existing therapies are aimed at suppressing inflammation rather than addressing the underlying biochemical processes. Thus, there remains a need in the art for materials and methods for treating fibrosis, including liver fibrosis.
Orf virus, the type species of the Parapoxvirus genus, causes localised proliferative skin lesions in ungulates and humans (Haig and Mercer, Vet Res 29, 311-26, 1998), with extravagantly proliferative and persistent lesions reported in immune-compromised individuals (Gurel et al., Eur J Dermatol 12, 183-5, 2002; Hunskaar, Br J Dermatol 114, 631-4, 1986; Savage et al., Proc R Soc Med 65, 766-8, 1972, Tan et al., Br J Plast Surg 44, 465-7, 1991). Orf virus infection initiates in the regenerating epidermis of wounded skin and the lesions progress through stages of erythema, papule, vesicle, pustule and then scab formation (Haig and Mercer, Vet Res 29, 311-26, 1998; Jenkinson et al., Vet Dermatol 1, 189-95, 1990). Orf virus lesions have been described as reminiscent of a sustained wound healing response, as they are characterized by extensive blood vessel proliferation and dilation and epidermal hyperplasia (Groves et al., J Am Acad Dermatol 12, 706-11, 1991, Savory et al., J Virol 74, 10699-706, 2000).
Vascular endothelial growth factors (VEGFs) are key regulators of angiogenesis during normal physiological and disease processes such as wound healing (Carmeliet and Jain, Nature 473, 298-307, 2011, Ferrara, Endocr Rev 25, 581-611, 2004, McColl et al., Apmis 112, 463-80, 2004). The VEGF family, which includes VEGF-A, VEGF-B, VEGF-C, VEGF-D and placental growth factor (PIGF), interact with the high-affinity VEGF receptors (VEGFRs), VEGFR-1, VEGFR-2 and VEGFR-3 (Koch et al., Biochem J 437, 169-83, 2011; Olsson et al., Nat Rev Mol Cell Biol, 7, 359-71, 2006). VEGF-A binds to both VEGFR-1 and VEGFR-2, whilst PIGF and VEGF-B bind exclusively to VEGFR-1. VEGF-C and VEGF-D interact with both VEGFR-2 and VEGFR-3. VEGFs also bind the co-receptors neuropilin (NRP)-1 and NRP-2, which enhance binding to the VEGFRs (Soker et al., J Cell Biochem 85, 357-68, 2002; Vadasz et al., Autoimmun Rev 9, 825-829, 2010). VEGF-A has been shown to promote angiogenesis by stimulating endothelial cell proliferation, migration and survival and promoting vascular permeability, primarily through VEGFR-2 (Holmes et al., Cell Signal 19, 2003-2012, 2007). VEGFR-1, however, appears to play a role in endothelial cell differentiation and migration, possibly by acting as a ligand-binding molecule, sequestering VEGF-A from VEGFR-2 signaling (Shibuya, Angiogenesis 9, 225-230, 2006). During cutaneous tissue repair, VEGF-A is highly expressed by keratinocytes and stimulates the formation of new blood vessels in the wound bed, supplying nutrients and oxygen needed for regeneration of the skin (Barrientos et al., Wound Repair Regen 16, 585-601, 2008; Brown et al., J Exp Med 176, 1375-1379, 1992; Nissen et al., Am J Pathol 152, 1445-1452, 1998). In addition, a number of studies have shown that VEGF-A also enhances healing by promoting re-epithelialization of wounds (Brem et al., J Invest Dermatol 129, 2275-2287, 2009; Li et al., Diabetes 56, 656-665, 2007; Michaels et al., Wound Repair Regen 13, 506-512, 2005; Romano Di Peppe et al., Gene Ther 9, 1271-1277, 2002). VEGF-A also increases vascular leakage and promotes the formation of disorganized blood vessels (Carmeliet, Nat Med 6, 1102-1103, 2000). Several other skin disorders are linked to a high presence of VEGF-A, such as psoriasis (Detmar, J Invest Dermatol 122, xiv-xv, 2004), skin cancer (Weninger et al., Lab Invest 75, 647-657, 1996), dermatitis herpetiformis and erythema multiforme (Brown et al., J Immunol 154, 2801-2807, 1995). VEGF-A overexpression in transgenic mice, with epidermal trauma, induces psoriatic-like lesions characterized by prominent angiogenesis, inflammation and epidermal hyperplasia (Canavese et al., Histol Histopathol 26, 285-296, 2011; Elias et al., Am J Pathol 173, 689-699, 2008; Xia et al., Blood 102, 161-168, 2003).
It has been reported that the extensive vascular changes found beneath the orf virus lesions are in part, if not solely due to the expression of a VEGF homolog encoded by this virus. In the absence of a functional viral VEGF, the infected lesions lack not only the striking proliferation of blood vessels and dermal edema but also the distinctive pattern of epidermal hyperplasia and rete ridge formation seen in wild-type infections (Savory et al., J Virol 74, 10699-10706, 2000; Wise et al., Virus Res 128, 115-125, 2007). It has also been reported that purified orf virus VEGF, which has been designated VEGF-E, promotes angiogenesis and epidermal regeneration through its interaction with VEGFR-2, but shows negligible vascular leakage and tissue inflammation as it fails to bind VEGFR-1 (Inder et al., Febs J 275, 207-217, 2008; Inoue et al., Arterioscler Thromb Vasc Biol 27, 99-105, 2007; Kiba et al., Biochem Biophys Res Commun 301, 371-377, 2003; Wise et al., J Biol Chem 278, 38004-38014, 2003; Wise et al., Cellular Microbiology 14 (9) 1376-1390, 2012; Zheng et al., Arterioscler Thromb Vasc Biol 26, 2019-2026, 2006; Zheng et al., Arterioscler Thromb Vasc Biol 27, 503-511, 2007).
Interleukin-10 (IL-10), also known as human cytokine synthesis inhibitory factor (CSIF), is an anti-inflammatory cytokine. In humans IL-10 is encoded by the IL10 gene. This cytokine is produced primarily by monocytes and to a lesser extent by lymphocytes and keratinocytes. It has pleiotropic effects in immunoregulation and inflammation. It down-regulates the expression of Th1 cytokines, MHC class II antigens, and costimulatory molecules on macrophages. It also enhances B cell survival, proliferation, and antibody production. IL-10 can block NF-κB activity, and is involved in the regulation of the JAK-STAT signaling pathway. IL-10 is capable of inhibiting synthesis of pro-inflammatory cytokines such as IFN-γ, IL-2, IL-3, TNFα and GM-CSF made by cells such as macrophages and regulatory T-cells. It also displays a potent ability to suppress the antigen-presentation capacity of antigen presenting cells. However, it is also stimulatory towards certain T cells and mast cells and stimulates B cell maturation and antibody production. IL-10 is mainly expressed in monocytes and Type 2 T helper cells (TH2), mast cells, CD4+CD25+Foxp3+ regulatory T cells, and also in a certain subset of activated T cells and B cells. It is released by cytotoxic T-cells to inhibit the actions of NK cells during the immune response to viral infection.
Many viruses exploit the strategy of using homologs of cellular cytokines or cytokine receptors to shield virus-infected cells from immune defenses and enhance virus survival in the host. Human cytomegalovirus (HCMV) is a species-specific betaherpesvirus that infects a majority of the world's population. HCMV establishes and maintains a lifelong latent infection in primitive myeloid lineage cells. Following terminal cell differentiation of these cells into myeloid dendritic cells (DCs) and macrophages, latent virus has the ability to reactivate, resulting in the production of new, infectious virions and often severe disease in immunocompromised individuals. Only a subset of viral genes are transcriptionally active during latency, including HCMV UL111A, a gene that encodes homologs of the potent immunomodulatory cytokine human interleukin-10 (hIL-10). UL111A is transcriptionally active during both productive and latent phases of infection and encodes several viral IL-10 proteins which exert a diverse range of immunomodulatory functions, including inhibition of cytokine synthesis and major histocompatibility complex (MHC) expression by myeloid cells, stimulation of B cells, and suppression of DC maturation and cytotrophoblast function. See Avdic, S, et al., Viral Interleukin-10 Expressed by Human Cytomegalovirus during the Latent Phase of Infection Modulates Latently Infected Myeloid Cell Differentiation, J. Virol. July 2011 85: 7465-7471. A number of herpes viruses also harbor homologs of IL-10. Epstein-Barr virus (EBV)-encoded IL-10 (ebvIL-10), the first viral homolog of IL-10 identified, shares many but not all of the biological activities of cellular IL-10 and may play an important role in the host-virus interaction. In addition to EBV, the Orf poxvirus (OV), which can infect humans, has its own IL-10 homolog, ovIL-10. See Kotenko, S V, et al., Human cytomegalovirus harbors its own unique IL-10 homolog (cmvIL-10), PNAS 97: 1695-1700 (2000). ORFV-IL-10 is functionally similar to cellular IL-10 in that it has the capacity to inhibit cytokine synthesis in human, ovine and murine monocytes (Wise et. al., J Gen Virol 88, 1677-1682, 2007; Fleming et al., J Virol 71, 4857-4861, 1997; Haig et al., Virus Res 88, 3-16, 2002; Imlach et al., J Gen Virol 83, 1049-1058, 2002), impairs the maturation of murine and human dendritic cells (Chan et al., J Gen Virol 87, 3177-3181, 2006, Lateef et al., J Gen Virol 84, 1101-1109, 2003,) but also costimulates mast cells and thymocytes (Fleming et al., J Virol 71, 4857-4861, 1997; Haig et al., Virus Res 88, 3-16, 2002; Imlach et al., J Gen Virol 83, 1049-1058, 2002).
Despite advances in the understanding of the principles underlying the wound healing process, there remains a significant unmet need for suitable therapeutic options for wound care and tissue repair and improving and/or promoting wound healing, including wounds that do not heal at expected rates, such as delayed-healing wounds, incompletely healing wounds, and compromised wound healing such as is seen in chronic wounds, scarring and abnormal or excessive scarring, including keloid and hypertrophic scarring, atropic scarring, widespread scarring, and scar contractures, as well as adhesions including surgical adhesions. There is a need in the art for improved methods and compositions for treating conditions such as those caused by acute and chronic wounds, inflammation, fibrosis, scarring, and adhesions.